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Shaoqing He, Hai-Ying Ma, Thomas Yorio; Astrocytes Promote Synapse Formation and Survival of Retinal Ganglion Cells at Early Phase During Endothelin Treatment. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5291. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Endothelin-1(ET-1) and its receptors are involved in the etiology of glaucoma. Previously, we reported that ET-1 treatment induced reactivation of astrocytes (ASTs) and apoptosis of retinal ganglion cells (RGC). Recently, it is reported that two newly-identified AST subtypes exhibit the neurotoxic and neuroprotective effect to RGCs. However, ET-mediated subtype changes of ASTs and their effect on RGC survival are largely unknown. This study aimed to studying the roles of ET-1 in the interaction between RGCs and AST subtypes.
Primary rat RGCs were isolated from rat pup retinas and ASTs from optic nerve. ASTs were seeded directly into RGC culture or seeded into a filter insert over RGC culture. Both co-cultures were treated with 100nM endothelin-1 for 24-72 hours and subsequent proteins were detected using immunocytochemistry. ET-1-mediated intracellular calcium was monitored in RGCs, ASTs and co-culture of RGCs and ASTs using Fura-2 AM calcium imaging. Cell death and survival were detected using LIVE/DEAD assay (calcein AM and ethidium homodimer-1 staining).
ET-1-induced elevation of [Ca2+]i was significantly attenuated in co-culture of RGCs and ASTs, and accordingly less cell death was also observed in both co-culture systems. More synapse formation was detected by immunostaining of Bassoon and Homer-1 in RGC-AST co-cultures. The longer co-culture time produced more synapses. In addition, co-culture attenuated the upregulation of phospho-JNK and ERK1/2 seen in RGC culture. However, ET-1 treatment didn’t increase synapse formation. ET-1 treatment or c-Jun overexpression in ASTs changes the gene expression profile in ASTs, and such changes may switch astrocyte subtypes.
ASTs co-cultured with RGCs not only stimulate more synapse formation in RGCs, but also decrease RGC death. The secreted factors from the activated AST may provide a neuroprotective role. c-Jun/AP-1 could be a crucial factor regulating the genes which are associated with AST subtypes. ET-1 induced the activation of astrocytes through a mechanism that may be involved in the control of calcium-mediated signaling and JNK/c-Jun pathway. The results suggest that the reactivation of ASTs initially serves to be neuroprotective; however, long-term it could lead to dysfunction in the optic nerve and affect RGC survival as astrocyte subtype functions change.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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